RU54196U1 - ANALYZER FOR DETERMINING THE TOTAL ANTIOXIDANT ACTIVITY OF OBJECTS - Google Patents

Abstract

The utility model relates to the field of physical and analytical chemistry and can be used to determine the total antioxidant activity of biologically active substances by voltammetry using a three-electrode cell. The essence of the utility model is that the measurement is based on the model reaction of oxygen electroreduction, which proceeds by a mechanism similar to the reduction of oxygen in human tissues and cells and is the main oxidative process in all objects of artificial and natural origin. The analyzer has high sensitivity, allows you to automatically measure, applicable for various objects of food, cosmetic and pharmaceutical industries and medicine.

Description

The utility model relates to the field of analytical instrumentation. In particular, to instruments of voltammetric analysis, it can also be used in the analysis of the properties of biologically active substances.

A known installation for analysis of mixtures containing a container with a solvent, a capillary tube, a flow sensor, a sample input device, a chromatographic column, a detector, an output signal raiser and a recording device. When the eluent flows through the flow sensor, an output signal of the flow sensor is generated at its output, which is determined by the flow of the eluent through a thermostatically controlled highly efficient pre-column of the flow sensor and has a linear dependence on the flow of the eluent. Thermostating of the pre-column in such a flow sensor is used to eliminate the dependence of the output signal of the flow sensor on changes in the ambient temperature (RU 2148824, 05/10/2000)

However, the known installation is not intended to measure the total antioxidant activity of biologically active substances.

A setup is described for determining the total antioxidant activity of biologically active substances by their electrochemical oxidation, which contains a solvent tank, a pump, a dispenser, made in Side of a multi-way valve connected to the pump outlet, with an analyte input device and from a thermostated electrochemical cell of an amperometric detector, an amplifier an electric signal, an analog-to-digital converter and an output signal recording device, the installation contains amperometric detector, and the electrochemical cell does not contain a reference electrode (RU 2238555 C1 07.25.2003).

The main disadvantage of this device is the complex and expensive equipment is not always justified for routine analyzes.

A known installation for determining the antioxidant activity of preparations containing a thermostatically controlled container with an oxidation substrate - polyunsaturated fatty acids of corn oil, where an antioxidant preparation is introduced and exposed to oxidation indicators, the container is connected to a spectrophotometer or to a chemiluminescent device equipped with a specialized computer

program, while antioxidant activity is inversely proportional to the level of lipid peroxidation products. The installation allows you to standardize the evaluation of the results, as well as use an available oxidation substrate (RU 2182706 05/20/2002)

This setup cannot be used to measure the total antioxidant activity of biologically active substances, it is suitable only for the analysis of individual substances.

The closest in technical essence to the proposed voltammetric analyzer is the voltammetric analyzer "TA2" (RU 2129713 C1 04/27/1999), the voltammetric analyzer contains a polarizing voltage generating unit connected to an electrochemical cell with electrodes, an indicator electrode, which is connected mechanically to the output of the electrochemical solution mixing device cells and electrically with the input of the measuring circuit connected respectively to the input and output of the control unit and you data water. The voltammetric analyzer is characterized in that a deactivation device for interfering with the analysis of dissolved substances is introduced into it, connected to one electrochemical cell and forming with it a single unit of the electrochemical sensor, controlled by signals from the control and data output unit. Device for mixing a solution of an electrochemical cell.

However, this device is designed to determine the concentrations of any elements determined by voltammetry. For this purpose, additional schemes have been introduced into it, although they allow one to increase the level of the analytical signal when determining certain elements (UV irradiation, inert gas sparging, ozonation), but they are far from being demanded (especially during mass routine analyzes). At the same time, the price of analyzes increases significantly. In addition, universal analyzers work only under the control of a personal computer, which greatly facilitates the work of the operator, but it is not possible to fully automate the processing of the analytical signal because of the difficulty in creating a single algorithm for processing many different analytical signals, therefore, the intervention of a qualified analytical chemist on these stages needed.

The objective of this utility model is to create an express, experimentally simple, cheap voltammetric device,

designed to determine the total antioxidant activity of biologically active substances, the procedure for processing the analytical signal and calculating concentrations is fully automatic and does not require operator intervention.

The problem is solved in that the voltammetric analyzer for determining the total antioxidant activity of biologically active substances contains a control and data output unit, a unit for generating polarizing voltage, an electrochemical cell with electrodes, a measuring transducer circuit. The control unit contains a personal computer connected to a microcontroller associated with a digital-to-analog converter (DAC) and an analog-to-digital converter (ADC). The electrochemical cell contains three electrodes: the working electrode is a mercury-film electrode, the auxiliary one and comparisons are silver chloride.

Figure 1 presents a schematic diagram of a utility model:

1. personal computer

2. microcontroller

3. An analog-to-digital converter (ADC)

4. Digital-to-analog converter (DAC)

5. Potentiostat

6. Auxiliary electrode

7. The reference electrode

8. Working electrode

9. Electrochemical cell

10. Transmitter

11. Magnetic stirrer

The voltammetric analyzer contains a control and data output unit, which includes a personal computer 1 connected via a microcontroller 2, the output of which is connected to an analog-to-digital converter (ADC) 3 and a digital-to-analog converter (DAC) 4. The DAC is the output of the control unit and data output, and the input of the ADC 3 is the input of the control unit and data output. The output of the DAC is connected to the polarizing voltage generating unit containing a potentiostat 5, the non-inverting input of which serves as the input to the polarizing voltage generating unit, and the output is connected to the auxiliary electrode 6. In this case, the inverting input of the potentiostat 5 is connected to the electrode

comparing 7 electrochemical cells. The working electrode 8 of the electrochemical cell 9 is connected to the input to the circuit of the measuring transducer 10, the output and the control input, which are connected respectively to the output of the control unit and data output, the outputs of which are connected with a magnetic stirrer 11.

The analyzer for determining the total antioxidant activity of objects depicted in figure 1 works as follows.

Consider the work of the analyzer on the example of determining the antioxidant activity of ascorbic acid with a concentration of 0.0001 g / ml and rutin

In a cell made in the form of cups of optically transparent quartz (25 ml), 10 ml of a background solution based on the selected solvent is poured and installed in the analyzer cell.

Three electrodes are lowered into the prepared cell: working - mercury-film 8, reference electrode 7 and auxiliary - silver chloride 6.

From the personal computer 1, the corresponding program for determining the antioxidant activity of objects is loaded into the microcontroller 2.

First, the process of taking voltammograms of the current of oxygen electroreduction (EV O ₂ ) (I ₀ ) in the absence of an antioxidant in the solution (in the "Background electrolyte" tab) is performed. The process is presented as follows. The solution is mixed for 10 seconds, then it is kept for 20 seconds in a calm state. After that, a linearly varying voltage begins to be applied to the electrodes in the range: from 0 V to - 1 V for a time of 40 seconds. The registration process is repeated 6 times. The device captures the wave of the OV O ₂ (I ₀ ), automatically digitizes it, finds the maximum current of the OV O ₂ and the potential at which the maximum of the current of the OV O ₂ is observed.

The program averages the obtained voltammograms of the current of the electromagnetic field O ₂ (I ₀ ) and displays the average curve on the display sheet. The panel provides data on the magnitude of the potential of the current peak and the maximum value of the current of the electromagnetic field O ₂ in the absence of antioxidant in the solution.

A solution with an antioxidant of 0.1 ml is added to the cell, and the process of taking voltammograms of the current of OV ₂ (I) current is started when an antioxidant is added to the solution. The cell turns on at the potential of the maximum current of the electric field O ₂ (I °) in the absence of an antioxidant in

solution. The solution is kept in this state for 180 seconds. Then again the waveform of the EV ₂ (I) waveform is taken. The registration process is repeated 6 times. The device captures the wave OV O ₂ (I), automatically digitizes it, finds the maximum current OV O ₂ .

Criteria are calculated by setting the necessary parameters. In the computer version, a graph of l-I / I ° is plotted against the time t of the experiment, which is displayed in the display sheet. This graph is processed by the least squares method and the slope of this graph is found. - d (l-I / I °) / dt.

K - kinetic criterion reflects the amount of active oxygen radicals that have reacted with an antioxidant (or total antioxidant content) per minute of time. Dimension: micromol / l • min.

Formula:

K = C ° _O2 • (lI / I °) / t,

where I is the current of oxygen electroreduction in the presence of an antioxidant in a solution, μA, I ° is the current of oxygen electroreduction in the absence of an antioxidant in a solution, μA, C ° _O2 is the initial oxygen concentration in a solution (values are given in the table, see the appendix), μg / l, t is the reaction time of the antioxidant with active oxygen radicals, min.

K * is the transformed kinetic criterion of antioxidant activity in relation to a standard antioxidant: it reflects the amount of absorbed active oxygen radicals when interacting with an antioxidant for a fixed time in relation to the standard. Dimension: dimensionless quantity.

Formula:

K ^* = K / K _Art .

where K is the value of the kinetic criterion for the test sample,

To _article - the value of the kinetic criterion for a substance selected as a standard (in this case, the values were calculated relative to dehydroquercetin)

IC _50% criterion showing at what concentration 50% inhibition of electroreduction of oxygen occurs in the presence of an antioxidant. Dimension: μmol / L.

The calculated data are shown in table 1.

An analyzer for determining the total antioxidant activity of objects Table 1. Substance name K, µmol / l • min K * relative to routine IC _50% , micromol / l routine 0.351 one 1,057 vitamin C 0.135 0.385 0.103

Claims (1)

An analyzer for determining the total antioxidant activity of objects, containing a control unit and data output, which includes a personal computer connected via a microcontroller, the output of which is connected to a digital-to-analog converter (DAC) and an analog-to-digital converter (ADC), a unit for generating polarizing voltage , an electrochemical cell with electrodes, a circuit of the measuring transducer, characterized in that the DAC is the output of the control unit and data output, and the input of the ADC is the input of the unit As for control and data output, the DAC output is connected to a polarizing voltage generating unit containing a potentiostat, the non-inverting input of which serves as the input to the polarizing voltage generating unit, and the output is connected to an auxiliary electrode, while the inverting input of the potentiostat is connected to the reference electrode of the electrochemical cell, the working electrode is electrochemical the cell is connected to the input to the circuit of the measuring transducer, the output and control input of which are connected respectively to the output of the unit ION and the data input, the outputs of which are connected with the mixing device.